Immunogenic compositions containing bacterial outer membrane vesicles

Abstract

This invention relates to outer membrane vesicles (OMVs) from Gram-negative bacteria. The vesicles comprise heterologous proteins or immunogenic fragments thereof expressed as lipoproteins in their membrane. The OMVs of the invention are capable of eliciting an immune response to the heterologous protein or to a fragment thereof when administered to a mammal. Other aspects of the invention relate to methods of preparing the OMVs and immunogenic compositions containing the same.

Description

TECHNICAL FIELD[0001]This invention relates to vesicles from Gram-negative bacteria. The vesicles comprise heterologous proteins in their membrane expressed as lipoproteins. The vesicles are particularly useful in immunogenic compositions, e.g. vaccines.BACKGROUND ART[0002]Bacterial Lipoproteins and Lipidation[0003]Bacterial lipoproteins are a class of peripherally anchored membrane proteins, which play key roles in basic bacterial physiology as well as in pathogenic mechanisms such as adhesion, colonization, invasion and immune evasion.[0004]While in Gram-positive bacteria lipoproteins cross the membrane and remain attached on its external side through their lipid chains, in Gram-negative bacteria they can be found in three different cellular compartments: 1) attached to the periplasmic side of the inner membrane, 2) attached to the periplasmic side of the outer membrane, and 3) exposed on the surface of the outer membrane (OM). Lipoproteins are synthesized in the bacterial cytosol...

AI Technical Summary

Benefits of technology

This patent describes a method for making outer membrane vesicles (OMVs) with increased amounts of a heterologous protein, specifically Streptolysin O (Slo) from Streptococcus pyogenes (GAS). The method involves inactivating or deleting a gene that produces OmpA, one of the major outer membrane proteins of Escherichia coli. This mutation results in increased production of OMVs carrying Slo, which has been shown to have protective activity against GAS challenge in mice. The patent also describes other mutations or genetic modifications that can be used to increase the production of OMVs in other Gram negative bacteria. Overall, this patent provides a method for making OMVs with increased amounts of a heterologous protein that can be used as a vaccine candidate against GAS and other Gram negative bacteria.

Problems solved by technology

However, the contribution of the different OMV-associated lipoproteins to the immunostimulatory properties of OMVs has not been dissected and fully elucidated so far.

1. OMVs are amenable for large scale production—In general, the amount of OMVs released by Gram-negative bacteria when grown under laboratory conditions is too low to allow their exploitation in biotechnological applications.

Infect Dis Ther., 5, 89-112) its main drawback is that the detergent treatment favors bacterial cell lysis with the consequence that the OMV preparations are heavily contaminated with cytoplasmic proteins (Ferrari et al., (2006) Proteomics, 6, 1856-1866).

However, a universal system working for any protein antigen has not been described yet.

A strategy that is effective for one specific antigen in terms of level of expression and elicitation of immune responses can be inefficient with other antigens.

2. Second, one potential issue encountered in using OMVs in vaccine applications is the presence of lipopolysaccharide (LPS), an endotoxin known to be reactogenic both in animals and humans.

In conclusion, LPS plays a key role in stimulating innate immunity and promoting adaptive immunity but, at the same time, it is reactogenic and potentially toxic.

Images